Recent studies have shown that immunotherapeutic approaches can reproducibly mediate the regression of large metastatic cancers in patients. We have identified over four dozen new cancer antigens presented by a variety of tumor types. The identification of the molecular nature of cancer associated antigens has enabled the evaluation of active immunization (cancer vaccine) approaches to the treatment of humans with metastatic disease. We recently treated 440 patients with metastatic cancer treated in the Surgery Branch, NCI. A variety of approaches were explored including the use of immunogenic peptides, recombinant viruses, naked DNA and peptide pulsed dendritic cells. We have completed a study of 128 patients treated with peptide vaccines over the course of 48 weeks in the adjuvant setting. This prolonged immunization generated antigen reactive precursors at levels up to 20% of all CD8+ cells. It is not known, however, whether these high levels can prevent cancer recurrence in the adjuvant setting. A study in 56 patients with metastatic melanoma who received peptide immunization in conjunction with the administration of anti-CTLA4 monoclonal antibody (MDX-010) resulted in an overall objective regression rate of 13% including metastases that regressed in the lung, liver, brain, lymph nodes and subcutaneous sites. Autoimmune toxicity was seen that highly correlated with cancer regression (P=0.008). Similar results were obtained in the treatment of patients with metastatic renal cancer.We have completed a study in 35 patients with metastatic melanoma refractory to treatment with IL-2 who underwent lymphodepleting conditioning with cyclophosphamide and fludarabine followed by the infusion of autologous, tumor reactive, expanded tumor infiltrating lymphocytes (TIL) and IL-2. Eighteen of 35 patients (51%) experienced an objective clinical response including regression of metastases in lung, liver, lymph nodes, brain and subcutaneous sites. Tumor regression was highly correlated with the persistence of the administered cells (p2=0.001) which sometime reached as high as 80% of all circulating CD8+ cells. Thus cell transfer approaches can be highly effective for the treatment of patients with metastatic melanoma. We are currently conducting studies of intensified lymphodepletion using whole body irradiation prior to cell transfer as well as studies of the addition of vaccination to the treatment regimen.We have recently completed a study of the genetic modification of lymphocytes to improve their anti-cancer activity. Genes encoding anti-tumor T cell receptors were incorporated into retroviruses which were then used to convert normal patient lymphocytes into cells from anti-cancer activity. The adoptive transfer of these cells to patients has resulted in the objective clinical regression of cancer in some patients and represents a very important proof that the principle gene manipulation of lymphocytes can be used to develop effective gene therapies for the treatment of patients with cancer. These studies are currently ongoing utilizing higher affinity T cell receptors that recognize melanoma as well as common epithelial cancers.